A new class of proteins with specific affinity for calcium ions has recently been discovered. These proteins contain the unusual amino acid gamma-carboxy-glutamic acid (Gla) which is dependent on vitamin K for its biosynthesis. Gla residues participate in the binding of Ca2+ and endow these proteins with the characteristic property of Ca2+-mediated adsorption to phospholipid surfaces. It is now clear that Gla-proteins are not restricted to blood plasma, bone, and kidney. All tissues and animal cells appear capable of vitamin K-dependent Gla-protein biosynthesis, although at widely different levels. This proposal addresses fundamental questions regarding the biochemical and physiological functions of these cellular Gla-proteins. Furthermore, since these proteins are almost certainly altered by the vitamin K-antagonistic oral anticoagulants such as warfarin, they may serve as a target for the tumor-inhibiting, antimetastatic properties of these drugs. Using several epithelial cell lines (RAG, mouse renal adenocarcinoma; MDCK, canine kidney) and primary kidney cells propagated in vitro, the requirements of vitamin K for growth, adhesiveness, and cellular locomotion will be explored by depletion studies and warfarin treatment. The microanatomical localization of cellular Gla-proteins will be revealed by autoradiography, after specific tritium labeling of Gla residues by a novel exchange-decarboxylation procedure. Labeling Gla by microsomal carboxylation in the presence of vitamin K and 14[CO2] provides a specific marker for purfication. The purified Gla-protein will be characterized and used to develop a specific radioimmunoassay. The involvement of this protein in metastasis of primary tumors is initially approached by studying its biosynthesis in primary and metastatic tumor cells from vitamin K-deficient and anticoagulated mice. The cellular Gla-protein may provide a new target for the rational design of chemotherapeutic agents and regimens based on vitamin K and its metabolites.